Alterations in gut microbiota composition are found in chronic inflammatory non-infectious and infectious diseases including chronic HIV infection. Such changes may influence host immune responses to pathogens and vaccines. However, our knowledge is limited regarding the influence of the baseline gut microbiome on the host immune responses to vaccines and pathogens. The proposed R01 application will utilize the nonhuman primate simian immunodeficiency virus (SIV) model of AIDS to investigate the influence of the gut microbiome on the induction of host immune responses to a SIV vaccine and to mucosal SIV challenge. This study will capitalize on the unique resource at the California National Primate Center (CNPRC) that includes specific pathogen free (SPF) and conventionally raised (non-SPF) rhesus macaques. Our preliminary data show that SPF and non-SPF macaques show distinct gut microbial communities that correlate with phenotypically and functionally diverse T and B cell subsets. SPF animals are raised free of commonly found chronic viral infections. In contrast, non-SPF animals are naturally exposed to chronic viral infections from other animals and are typically positive for multiple herpes viruses. SPF and non-SPF macaque cohorts permit investigation of the influence by altered microbiota associated with, and in the context of chronic subclinical infection induced alterations of immune responsiveness to SIV vaccines. This opportunity is highly relevant to understanding the variability in vaccine responses among populations from developing and developed worlds that have different levels of pre-existing chronic viral infections and potentially altered gut microbiomes. We will utilize a novel vaccine formulation based on RhCMV vectors expressing SIV antigens (RhCMV-Gag, RhCMV-Retanef, RhCMV-Env) and an RhCMV vector expressing an SIV antigen fused to a TLR5 ligand (RhCMV-SIV-Gag-FliC) to determine vaccine immunogenicity and efficacy in the context of distinct gut microbiota composition through the use of the unique SPF and non-SPF cohorts. We will test the hypothesis that gut microbiota associated with subclinical viral infections typical to many human populations may impact the magnitude and diversity of the immune responses to a SIV vaccine and vaccine efficacy. Aim 1: Investigate the impact of distinct gut microbiome composition and diversity on the SIV vaccine-induced immune responses of SPF and non-SPF rhesus macaques. Rhesus macaques will be immunized with a novel RhCMV-SIV-based vaccine that contains a TLR5 adjuvant and innate and virus-specific cellular and humoral immune responses will be compared between SPF and non-SPF groups. Aim 2: Investigate the impact of the gut microbiome composition and diversity on the host immune responses to SIV infection in immunized and naive, SPF and non-SPF rhesus macaques. The proposed study has potential to provide insights into understanding the influence of baseline microbiome composition and associated immune cell activation status on the host immune responses to HIV vaccines and the virus and help develop innovative approaches to therapies.